CONICET | Buscador de Institutos y Recursos Humanos

Aldolases became widely used enzymes due to their ability to catalyze the stereoselective C-C bond formation through a condensation reaction that involves carbonylic compounds, known as aldolic condensation . In particular, these enzymes are useful in the synthesis of carbohydrates , since these molecules contain multilpe chiral carbon atoms carrying hydroxyl groups of similar reactivity. Among them, 2-deoxyribose-5-phosphate aldolase (DERA) is the only aldolase that accepts two aldehydic substrates. The condensation reaction between D-glyceraldehyde-3-phosphate (G3P) and acetaldehyde catalyzed by DERA generates a chiral carbon with S configuration, producing 2-deoxyribose-5-phosphate (DR5P). As any aldolase, DERA shows high specificity towards the donor substrate, G3P. However, it tolerates other aldehydes as acceptor molecules. Besides acetaldehyde, it has been found that DERA is capable of accepting propanal, acetone and fluoroacetone, enabling the production of substituted 2-deoxyriboses. These compounds are important intermediates in the synthesis of carbohydrates as well as nucleosides . The latter have high comercial interest owing to the fact that they are involved in the preparation of antiviral and antisense drugs and deoxyribonucleotides (dNXPs) for PCR techniques. Both the chemical synthesis of deoxyriboses and the following production of deoxynucleosides (dN) require several and complex steps including those of protection and deprotection. For this reason, the study of simpler, cleaner and economic alternative paths becomes highly interesting. With the aim of selecting whole cells of microorganisms containing DERA activity useful to obtain DR5P, our bacteria collection was first screened for finding those bacteria capable to metabolize 2-deoxyribose as source of carbon and energy. After the secondary screening, microorganisms that tolerated high acetaldehyde concentration were detected. Finally, ten microorganisms belonging to the genera Bacillus, Xanthomona, Streptomyces and Erwinia were identified. These bacteria were tested for the DR5P production through the aldolic condensation described above. Among them, four microorganisms generated only 2-deoxyribose but only one produced both 2-deoxyribose and DR5P. In order to obtain DR5P in a more economical way , glucose was used as substrate to allow the in situ generation of G3P. Regarding the necessity of ATP, several cell poration techniques were developed, including not only the use of surfactants like NP-40, Tween-20 and Triton X-100, but also chelant agents as EDTA and solvents like xylene. Reaction products detection was conducted through thin layer chromatography (TLC). 2-deoxyribose and DR5P were quantified by colorimetric methods based on diphenylamine and cysteine chloride reactions

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